建立载脂蛋白E 基因敲除小鼠骨髓髓源性生长因子缺失模型及鉴定. (Chinese)

Establishment and identification of a myeloid-derived growth factor deficiency model in apolipoprotein E knockout mice. (English)

BACKGROUND: There are few studies on myeloid-derived growth factor (MYDGF) and atherosclerosis and metabolic diseases. Therefore, a mouse model of bone marrow specific MYDGF deficiency with apolipoprotein E (ApoE) knockout is particularly important for studying the relationship between MYDGF and atherosclerotic diseases. OBJECTIVE: To establish and identify the mouse model of bone marrow specific MYDGF deficiency with ApoE knockout. METHODS: ApoE^flox/flox mice and bone marrow specific MYDGF^flox/flox mice were purchased from the Shanghai Model Organisms Centre, Inc (Shanghai, China). Five ApoE^flox/flox male mice were selected to mate with 10 MYDGFflox/fiox female mice, and 30 F1 progeny mice with genotype of MYDGF^flox/+ApoE^flox/+ were obtained and mated with each other. Finally 30 mice with genotypes of MYDGF^flox/floxApoE^flox/flox and 34 ApoE^flox/flox progenies were obtained. The MYDGF^flox and ApoE^flox genotypes were identified by PCR, and the body length and body mass of MYDGF^flox/floxApoE^flox/flox mice and ApoE^flox/flox mice at 2 months were recorded. The expression of MYDGF protein in the mouse bone marrow tissues was detected by western blot assay, and immunofluorescence was used to detect the expression of MYDGF in the mouse bone marrow cells. The plaque area of the thoracic aorta in the mouse was detected by oil red O staining. The study was approved by the Animal Ethic Committee of PLA General Hospital of Central Theater Command. RESULTS AND CONCLUSION: There were 34 ApoE^flox/flox mice and 30 MYDGF^flox/floxApoE^flox/flox mice, with no significant difference in their body length and body mass (both P > 0.05). The relative expression level of MYDGF protein in the bone marrow of MYDGF^flox/floxApoE^flox/flox mice was significantly lower than that of ApoE^flox/flox mice (P < 0.05). The percentage of plaque area in the thoracic aorta of MYDGF^flox/floxApoE^flox/flox mice was significantly higher than that of ApoE^flox/flox mice (P < 0.05). Therefore, MYDGF gene was successfully knocked out in ApoE^flox/flox mice, and the homozygous mouse model of bone marrow knockout MYDGF gene in ApoE knockout mice was successfully constructed and identified by genotype and protein tissue levels. To conclude, MYDGF gene deficiency can aggravate atherosclerosis in ApoE knockout mice. [ABSTRACT FROM AUTHOR]

目前关于髓源性生长因子(myeloid-derived growth factor，MYDGF)与动脉粥样硬化及代谢性疾病的研究较少，因此建立一种载脂蛋白 E(apolipoprotein E，ApoE)敲除骨髓MYDGF缺失的小鼠模型对于研究MYDGF与动脉粥样硬化疾病的关系尤为重要。 目的：建立ApoE敲除骨髓MYDGF特异性缺失小鼠模型并进行鉴定。 方法：ApoE敲除小鼠和骨髓特异性MYDGF敲除小鼠，均购于上海南方模式生物科技股份有限公司。选取5只ApoE敲除小鼠与10只骨 髓特异性MYDGF敲除小鼠进行交配，得到基因型为MYDGFflox/+ApoE^flox/+ F1子代小鼠30只，MYDGF^flox/+ApoE^flox/+互相进行交配，一共得到 基因型为MYDGFflox/floxApoEflox/flox双敲除小鼠30只和ApoE^flox/flox小鼠子代小鼠34只。采用PCR法进行MYDGF^flox及ApoE^flox基因型鉴定，记录 MYDGF^flox/floxApoE^flox/flox双敲除小鼠与ApoE^flox/flox小鼠2月龄的体长及体质量，采用Western blotting和免疫荧光检测2组小鼠骨髓组织MYDGF蛋白 相对表达量，油红O染色检测2组小鼠胸主动脉斑块面积。实验方案经中国人民解放军中部战区总医院动物实验伦理委员会批准。 结果与结论：①得到基因型为ApoE^flox/flox小鼠共34只、MYDGF^flox/floxApoE^flox/flox小鼠共30只，2组小鼠体长、体质量比较差异均无显著性意义(均 P > 0.05)；②MYDGF^flox/floxApoE^flox/flox小鼠骨髓组织MYDGF蛋白相对表达量显著低于ApoE^flox/flox小鼠(P < 0.05)；③MYDGF^flox/floxApoE^flox/flox小鼠胸主 动脉的斑块面积百分比显著大于ApoE^flox/flox小鼠(P < 0.05) ；④结果证实成功敲除了ApoE小鼠的MYDGF基因，说明成功构建ApoE敲除骨髓特 异性MYDGF缺失小鼠模型，并经基因型及蛋白组织水平鉴定；MYDGF基因缺乏可以加重ApoE基因敲除小鼠的动脉粥样硬化。 [ABSTRACT FROM AUTHOR]

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